Published September 1, 1929
THE EFFECT OF CATAPHORESIS ON POLIOMYELITIS
VIRUS
BY P. K. OLITSKY, M.D., C. P. RHOADS, M.D., AND P. H. LONG, M.D.
(From the Laboratories of The Rockefeller Institute for Medical Research)
(Received for publication, May 25, 1929)
Method
The apparatus used was a modification of that described by Field and Teague
(6) and by Todd (1). The details will be given by Olitsky and Long (4) in another
communication. It is sufficient to say here that three large U tubes were employed, the combined capacity of Which was about 96 cc. of fluid. The ends of
the connecting tubes which dipped into the fluid under observation contained I
per cent agar and 1 per cent NaC1. This furnished a convenient method for c o l
lecting the particles containing th e virus, or the virus itself, which migrated with
the current. From 114 to 117.5 volts and 0.8 to 4.2 milliamperes were passed for
3 hours through the fluid under examination between non-polarizable electrodes.
273
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Recent investigations in the field of filterable viruses have been
marked by a tendency to apply to the disease-producing agents of this
class certain biological principles which govern the reactions of bacteria. Of particular interest have been the obsercations on the behavior of viruses in an electrical field (1, 2, 3). By means of cataphoresis of virus suspensions, it has been found that the effective
agent could be revealed in tissues which, by the usual tests of animal
inoculation, failed to show activity. This results from the migration
of the virus, or possibly, of the material in a suspension carrying the
virus, to one or the other p01e, depending on the charge. Thus vaccine virus under ordinary conditions of hydrogen ion concentration
migrates to the anode (2, 3). By this method, furthermore, vaccine
virus has been isolated from rabbits wholly recovered from infection
and several weeks or months after vaccination (3, 4). Finally, cataphoresis has been of use in separating vaccine virus from neutral virusimmune serum mixtures (5). Since these observations have a bearing on immunity in virus diseases, it was determined to ascertain the
behavior of the poliomyelitic virus in an electrical field.
Published September 1, 1929
274
E F F E C T OF CATAPHORESIS O N POLIOMYELITIS VIRUS
At the end of the period the agar plugs were removed under sterile conditions,
ground in 1 cc. of Ringer's solution, and inoculated intracerebrally in monkeys,t
The mode of preparation of the fluid will be given with each of the following
experiments.
T h e first step was to determine whether the virus of poliomyelitis
possessed the p r o p e r t y of m i g r a t i n g in an electric field.
F r o m the fact t h a t the a n i m a l inoculated with anodic m a t e r i a l
developed poliomyelitis f i r ~ , we were led t o suppose t h a t the virus
p r o b a b l y carried a negative charge. Since the m o n k e y inoculated
with the cathodic m a t e r i a l also c o n t r a c t e d the disease, although a f t e r
a s o m e w h a t longer incubation period, it was t h o u g h t t h a t p r o b a b l y
the virus suspension was too strong, a n d p a r t of the virus a d h e r e d to
the cathodic agar. T o obviate this difficulty we n e x t e m p l o y e d a
B e r k e f e l d " V " filtrate of a 5 p e r cent suspension of the active n e r v o u s
tissue.
Experiment 2.--The spinal cord from a monkey with poliomyelitis was made
into a 5 per cent suspension in distilled water. The emulsion was passed through
a Berkefeld "V" candle and placed in the cataphoresis apparatus. The conditions
x All experimental procedures on animals were made with the aid of deep ether
anesthesia.
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Experiment 1.--A piece of spinal cord of a monkey dead of typical experimental
poliomyelitis was used. The nervous tissue was emulsified in 100 cc. of distilled
water under aseptic conditions. The cataphoresis test was done as follows: milliamperes current, 0.8; potential drop, 117 volts; time, 3 hours; pH of the suspension, 6.9. After 30 minutes the fluid at the cathodes was dear and that at the
anodes cloudy. After 2 hours the cathodic material was quite clear for a distance
of 2 cm. from the surface and the remainder was partly clear. The agar from the
positive poles contained a diffuse whitish cloud and that from the negative pole
was translucent. Macacus rhesus 1 was inoculated intracerebrally with 1 cc. of
Ringer's solution containing the ground agar from the positive poles. On the
ninth day after inoculation the animal was excited and tremulous and both legs
were paralyzed. On the tenth day the prostrate monkey was etherized. The
gross and microscopical lesions of the spinal cord and brain were typical of poliomyelitis. Macacus rhesus 2 received a similar intracerebral inoculation of Ringer's
solution containing the agar from the negative poles of the same experiment. On
the ~teenth day after inoculation all four limbs were paralyzed and the animal was
prostrate. The monkey was etherized and the gross and microscopic lesions of the
nervous system were typical of poliomyelitis.
Published September 1, 1929
P. If. OLITSKY, C. P. RIIOADS, A N D
P. H. L O N G
275
were: milliampere.s, 0.8; potential drop, 116 volts; time, 3 hours. The pH of the
filtrate was 6.5. Contrary to the previous experiment, in which suspension was
used, the cleating in this case appeared at the positive pole and the cathodic
material remained cloudy. Macacus rhesus 3 was inoculated intracerebrally with
1 cc. of Ringer's solution containing the ground agar from the positive pole. The
fifth day after inoculation the animal was excited, weak, and ataxic. The next
day a marked tremor and partial paralysis of all limbs were present. On the
seventh day the animal was prostrate, and on the eleventh it was etherized. The
lesions of the nervous system were typical of poliomyelitis. Macacus rhesus 4
(control) was inoculated the same day with the material from the negative pole.
This monkey remained well throughout the period of observation.
Experlr~ent 3.--Poliomyelitis material of the pooled mixed virus strain which
had been glycerolated for some time was made into a 5 per cent emulsion in salt
solution and passed through a Berkefeld "V" filter. 50 cc. of the filtrate was
mixed with an equal volume of human convalescent serum. 4 cc. of this mixture
was inoculated intraspinally and intracerebrally in Macacus rhesus 5 (control).
No symptoms developed. The material was therefore considered to have been
neutralized. The remainder of the mixture was subjected to cataphoresis under
the following conditions: milliamperes, 4.2; potential drop, 114 volts; time, 3
hours; pH = 6. Macacus rl~.sus 6 was inoculated intracerebrally with 1 cc. of
Ringer's solution containing the agar from the positive pole. On the eleventh day
after inoculation the animal was excited, slow in movement, and tremulous.
Ptosis and a right facial paralysis were present. On the twelfth day the monkey
was ataxic and partially paralyzed in all four limbs. The thirteenth day the prostrate animal was etherized. The gross and microscopic lesions in the spinal cord
and brain confirmed the diagnosis of poliomyelitis. Macacus rhesus 7 (control)
was inoculated intracerebrally on the same day with the emulsified agar from the
negative pole of the same experiment. No symptoms developed.
T h e experiment shows unmistakably t h a t active poliomyelitis
virus can be separated from a non-infective, neutral mixture of the
virus and its specific immune serum. W e can thus conclude t h a t the
virus is not killed b y the antiserum b u t only held in some sort of an
ineffective combination. T h e result also confirms the previous obser-
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I t will be n o t e d t h a t the virus collected at the positive pole. I t is of
interest t h a t the clouding due to migration of proteins in the fluid
appeared at t h e pole opg0site to the one at which the virus was found.
T h e next experiment was designed to recover b y cataphoresis the
virus from a non-infectious, neutral mixture of poliomyelitis virus a n d
convalescent serum.
Published September 1, 1929
276
E F F E C T OF CATAPHORESIS ON P O L I O M Y E L I T I S VIRUS
ration that under the conditions of the experiment poliomyelitis virus
bears a negative charge.
Since it was found possible to regain active poliomyelitis virus from
the neutral mixture, it was thought advisable to attempt to isolate
virus from an animal which had recovered from the disease.
This experiment brings o u t the interesting fact that active virus
persists in the nervous tissues for a period after recovery from poliomyelitis. Since ordinary inoculation of central nervous tissues before
cataphoresis was without effect, the positive result indicates either
that the antibody concentration in the tissues is sufficient to have
neutralized the virus associated with it on injection in our experiment,
or that in the recovered monkeys the virus is present in too minute
quantity to be detected by the usual mode of inoculation. The tests
described demonstrate that the virus of poliomyelitis may be separated in effective quantity from neutral mixtures of virus and immune
serum or further that it can be concentrated at the positive pole from
dilute virus filtrates.
CONCLUSIONS
1. Under ordinary conditions of hydrogen ion concentration the
virus of poliomyelitis, as such, or associated with particles in fine sus-
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Experiment 4.--A monkey, Macacus rhesus 8, was selected which had developed
typical poliomyelitis 6 days after an intracerebral inoculation of virus. Almost
complete paralysis of both arms and partial paralysis of the legs had existed for 4
days, followed by rapid recovery, so that by the eleventh day there was little
disability. During the next 12 days the monkey ran and jumped about with
almost normal vigor. At this time, or 23 days after infection, the animal was
ethcrized and the spinal cord made into a 5 per cent suspension in salt solution.
Cataphoresis was carried out under the following conditions: milliamperes, 0.8;
potential drop, 117.5 volts; time, 3 hours; pH = 6.8. Mac~us rhesus 9 was inoculated intracerebrally with the agar suspension from the positive pole. On the
eighteenth day tremor, ataxia, and partial paralysis of both arms appeared. On
the nineteenth the monkey was prostrate and was ether~ed. The gross and microscopic examination of the central nervous system confirmed the diagnosis of poliomyelitis. Mazacus rhesus 10 (control) was inoculated intracerebrally with the
material from the negative pole and 11 (control) with the suspension of central
nervous system before cataphoresis had been performed. Neither animal developed poliomyelitis.
Published September 1, 1929
P. K. OLITSKY, C. P. RIIOADS, A N D
P. H. L O N G
277
pension, migrates in an electrical field to the anode. It follows that
the virus bears an electronegative charge.
2. By means of cataphoresis, the virus can be recovered from a
non-infective mixture of virus and specific immune serum.
3. B y the same means it is possible to reveal the presence of virus
in the central nervous system of a monkey which has recovered from
the active stage of experimental poliomyelitis.
BIBLIOGRAPHY
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1. Todd, C., Brit. Your. Exp. Path., 1927, 8,369.
2. Douglas, S. R., and Smith, W., Brit. Your. Exp. Path., 1928, 9, 213.
3. Olitsky, P. K., and Long, P. H., Science, 1929, 69, t70.
4. Olitsky, P. K., and Long, P. H., 3. Exp. Med., 1929, 50, 263.
5. Long, P. H., and Olitsky, P. K., Science, 1929, 69, 72; a more detailed article
is in press.
6. Field, C. W., and Teague, O., Y. Exp. Med., 1907, 9, 86.